Marek's disease is a lymphoproliferative disease of chickens caused by Marek's disease virus (MDV). MDV, a naturally occurring herpesvirus, infects bursa-derived and thymus-derived lymphocytes in chickens, and may subsequently induce a lymphoma of thymus-derived lymphocytes. MDV is a designation of a family of avian herpesviruses. For example, MDV1 is a virulent strain of herpesvirus in chickens, MDV2 is a naturally attenuated herpesvirus strain in chickens, and MDV3 is a nonpathogenic herpesvirus of turkey.
Since Marek's disease is contagious, the virus has become an important pathogen of chickens, particularly in an environment of large scale breeding such as in the poultry industry. Currently, Marek's disease is controlled by vaccination of embryos at 17-19 days of incubation, or one day old chicks.
The application of recombinant DNA techniques to animal viruses in general has a recent history. The first viruses to be engineered have been those with the smallest genomes. For example, in the case of the papovaviruses, because these viruses are so small and cannot accommodate much extra DNA, their use in genetic engineering has been as defective replicons. Thus, foreign DNA sequence expression from these viruses requires a wild-type helper virus and is limited to cell culture systems. On the other hand, for adenoviruses, there is a small amount of nonessential DNA that can be replaced by foreign sequences. This technique has also been applied to portions of the herpesvirus genome in an avian herpesvirus (see U.S. Pat. No. 5,853,733 to Cochran et al).
The cases of deletion or insertion of genes into herpesviruses demonstrate that it is possible to genetically engineer herpesvirus genomes by recombinant DNA techniques. In the past, the methods that have been used to insert genes involve homologous recombination between the viral DNA cloned in plasmids and purified viral DNA transfected into the same animal cell. However, the extent to which one can generalize the location of the deletion and the sites for insertion of foreign DNA sequences is not known from these previous studies.
The identification of suitable DNA sequence insertions sites in avian herpesviruses are valuable for the development of new vaccines. The selection of (i) a suitable virus and (ii) the particular portion of the genome to use as an insertion site for creating a vector for foreign DNA sequence expression, however, pose a significant challenge. In particular, the insertion site must be non-essential for the viable replication of the virus, as well as its operation in tissue culture and in vivo. Moreover, the insertion site must be capable of accepting new genetic material, while ensuring that the virus continues to replicate.
What is needed is the identification of novel viruses and gene insertion sites for the creation of new viral vectors.